Assessing Heteroplasmic Load in Leber's Hereditary Optic Neuropathy Mutation 3460G→A/MT-ND1 with A Real-Time PCR Quantitative Approach Anna Genasetti,

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Assessing Heteroplasmic Load in Leber's Hereditary Optic Neuropathy Mutation 3460G→A/MT-ND1 with A Real-Time PCR Quantitative Approach Anna Genasetti, Maria L. Valentino, Valerio Carelli, Davide Vigetti, Manuela Viola, Evgenia G. Karousou, Gian Vico Melzi d'Eril, Giancarlo De Luca, Alberto Passi, Francesco Pallotti The Journal of Molecular Diagnostics Volume 9, Issue 4, Pages 538-545 (September 2007) DOI: 10.2353/jmoldx.2007.060183 Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 Pedigree of the 3460G→A/ND1 LHON family analyzed in this study. Proband position in the pedigree is indicated by an arrow. For all members, the position of the generation and family is indicated. The age of each member is reported at the time of blood sampling. The Journal of Molecular Diagnostics 2007 9, 538-545DOI: (10.2353/jmoldx.2007.060183) Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 2 Calibration curves of mtDNA amplification obtained by serial dilutions of plasmids using TaqMan MGB probe. A: Calibration curve of mutant mtDNA; B: calibration curve of wild-type mtDNA; and C: calibration curve of total mtDNA. Slopes of each calibration curve with the corresponding calculated efficiency and correlation coefficients (R2) are reported for each set of primers. The Journal of Molecular Diagnostics 2007 9, 538-545DOI: (10.2353/jmoldx.2007.060183) Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 3 Correlation of expected and observed percentages of 3460G→A mutant mtDNA by real-time ARMS-PCR analysis, using TaqMan assay, of 12 samples containing various proportions of mutant mtDNA. Equation for the line: y = 1.02x + 1.06%. The Journal of Molecular Diagnostics 2007 9, 538-545DOI: (10.2353/jmoldx.2007.060183) Copyright © 2007 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions